Many drugs (therapeutic agents) have undesirable properties, for instance, low oral drug absorption, toxicity, poor patient compliance etc., that may become pharmacological, pharmaceutical, or pharmacokinetic barriers in clinical drug application. Among the various approaches to minimize the undesirable drug properties, while retaining the desirable therapeutic activity, the chemical approach using drug derivatisation offers perhaps the highest flexibility and has been demonstrated as an important means of improving drug efficacy (Hyo-Kyung Han and Gordon L. Amidon AAPS PharmSci. 2000; 2 (1)).
The conventional approach that is adopted to minimize the toxic side effects associated with the therapeutic agents has been to derivatise one or more functional groups present in the drug molecule. The derivatives are then assessed for their therapeutic efficacy as well as toxicity. The carboxylic acid group is often present as an active functional group for derivatisation in several therapeutic agents. Non-steroidal anti-inflammatory drugs (NSAIDs) represent one of the best class of drugs containing a carboxylic acid group as an active functional group. NSAIDs are also the most commonly used drugs to relieve pain, symptoms of arthritis and soft tissue inflammation. Most patients with rheumatoid arthritis receive NSAIDs as a first-line treatment which is continued for prolonged periods. Although, NSAIDs provide anti-inflammatory and analgesic effects, they also have adverse effects on the upper gastrointestinal (GI) tract. The occurrence of GI toxicity appears to be strictly correlated to the mechanism of action of these drugs, namely the inhibition of the enzyme cyclooxygenase. In fact, inhibition of platelet cyclooxygenase, which causes prolonged bleeding time, and inhibition of cyclooxygenase in gastrointestinal mucosa, which results in a decreased synthesis of cytoprotective gastric prostaglandins, represent the major cause of serious gastrointestinal toxicity (Symposium on “New Anti-inflammatory agents: NO-NSAIDs and COX-2 inhibitors” part of the 11th international conference on “Advances in prostaglandin and leukotrine research: Basic science and new clinical applications” held in Florence (Italy), Jun. 4-8, 2000). This problem has been solved by derivatisation of carboxylic acid group of NSAIDs into its ester and amide derivatives.
Another common approach to minimize adverse effects of the known drugs or therapeutic agents consists of attaching a carrier group to the therapeutic agents to alter their physicochemical properties and then subsequent enzymatic or non-enzymatic cleavage to release the active drug molecule (therapeutic agent). The therapeutic agent is linked through a covalent linkage to specialized non-toxic protective groups or carriers or promoieties in a transient manner to alter or eliminate undesirable properties associated with the parent drug to produce a carrier-linked prodrug.
Indeed, a more recent strategy for devising a gastric-sparing NSAID involves chemically coupling a nitric oxide (NO) releasing moiety to the parent NSAID. The approach and possibility of combining a few classes of drugs bearing different functional groups susceptible of derivatisation with NO-donating moieties has been described by Menlo Bolla et al., in Curr. Topics Med. Chem. 2005; 5: 707-720.
Nitric oxide is one of the most important mediators of mucosal defense, influencing factors such as mucus secretion, mucosal blood flow, ulcer repair and the activity of a variety of mucosal immunocytes (Med Inflammation, 1995; 4: 397-405). It has been reported to play a critical role in maintaining the integrity of the gastroduodenal mucosa and exerts many of the same effects as endogenous prostaglandins (Drugs Fut 2001; 26(5): 485). Several mechanisms are considered to underlie its protective effect in the stomach including vasodilation of local mucosal blood vessels, inhibition of leukocyte adhesion and inhibition of caspase enzyme activity. The inactivation of caspase(s) appears to be an important factor in the GI tolerance of nitric oxide releasing NSAIDs (NO-NSAIDs) (J. E. Keeble and P. K. Moore, British Journal of Pharmacology, 2002; 137: 295-310). Nitric oxide can thus be used to devise a gastric-sparing NSAID. Compounds that release nitric oxide in small amounts over a prolonged period of time may be very useful for the prevention of gastrointestinal injury associated with shock and with the use of drugs that have ulcerogenic effects (Muscara M. N.; Wallace J. L. American Journal of Physiology, Gastrointestinal and liver physiology, 1999; 39: G1313-1316).
In recent years, several NO-releasing non-steroidal anti-inflammatory drugs (NO-NSAIDs) have been synthesized by an ester linkage formed through coupling of a NO-releasing chemical spacer group to the carboxylic acid moiety of a conventional NSAID. The use of various aliphatic, aromatic or heterocyclic chemical spacers makes it possible to alter various physicochemical properties and kinetics of nitric oxide release (Berguad et al., Ann. N.Y. Acad. Sci. 1962: 360-371 (2002)). The first NO-asprin drug NCX 4016, which was synthesized relatively recently, consists of an aspirin molecule linked by an ester bond to a molecular spacer, which in turn, is linked to a nitro-oxy ester group (Dig Liver Dis 2003; 35 (suppl. 2):9-19). A number of NO-NSAID hybrid compounds, namely NO-naproxen (Naproxcinod), NO-flurbiprofen (HCT 1026), NO-ibuprofen, NO-diclofenac and NO-indomethacin have been disclosed in the patent numbers EP 722434B1, U.S. Pat. No. 6,613,784 B1 and U.S. Pat. No. 7,220,749 B2, respectively. European Patent EP 722434B1 discloses nitrate esters of the derivatives of propionic acid, 1-(p-chlorobenzoyl)-5-methoxy-2-methyl-3-indolylacetic acid and 5-benzoyl-1,2-dihydro-3H-pyrrolo[1,2-a]pyrrole-1-carboxylic acid having anti-inflammatory and/or analgesic activity. U.S. Pat. No. 6,613,784 B1 discloses nitro derivatives of NSAIDs, for instance, flurbiprofen, indomethacin, aspirin, naproxen and diclofenac. U.S. Pat. No. 7,220,749 B2 discloses novel nitrosated and/or nitrosylated derivatives of COX-2 selective inhibitors. U.S. Patent Application Publication no. 20080293781A1 describes O-acyl salicylic acid derivatives bearing a NO donor moiety. U.S. Pat. No. 7,199,154 B2 discloses nitrosated or nitrosylated prodrugs for COX-2 selective inhibitors that are useful for treating COX-2 mediated diseases or conditions and which can be administered alone or in combination with low-dose aspirin. The compounds are effective in treating chronic COX-2 mediated diseases or conditions, reducing the risk of thrombotic cardiovascular events and possibly renal side effects and at the same time reduce the risk of GI ulceration and bleeding. US Patent Application Publication no. 20060058363 A1 discloses nitric-oxide releasing prodrugs of celebrex and valdecoxib which are useful in the treatment of COX-2 mediated diseases. The compounds may be used as a combination therapy with low-dose aspirin to treat COX-2 mediated diseases or conditions while simultaneously reducing the risk of thrombotic cardiovascular events.
Nitric oxide (NO) also plays an important role in numerous other physiological and pathophysiological conditions, e.g. blood pressure regulation, inflammation, infection and the onset and progression of malignant and cardiovascular diseases (Lirk, P., Hoffmann, G., and Rieder, J. Curr. Drug Targets Inflamm. Allergy 2002; 1:89-108). Though delivery of supplementary NO in the form of NO-donor drugs has long been an attractive therapeutic strategy (Ian L Megson, David J Webb, Expert Opin. Investing. Drugs, 2002; 11(5): 587-601), in recent years, with the advent of NO-NSAID approach and because of the beneficial biochemical and pharmacological properties of nitric oxide, the strategy of linking NO-releasing moieties has been extended to a wide array of therapeutic agents selected from cardiovascular drugs, for instance, Angiotensin converting enzyme (ACE) inhibitors, calcium antagonists and beta-blockers, antitumor agents, antihistamines, glucocorticoids, etc. The aim of this strategy is to synthesize prodrugs that retain the pharmacological activity of the parent drug molecule coupled with the benefits of the biological actions of NO in reducing the adverse effects of the parent drug molecule.
U.S. Pat. Nos. 6,610,676 and 7,524,836 B2 disclose nitrate esters and nitrooxy derivatives of steroidal compounds having anti-inflammatory, immunodepressive and angiostatic activity or gastrointestinal activity.
PCT Application Publication WO2007099548A1 discloses 11β-hydroxyandrosta-4-3-one compounds which possess useful anti-inflammatory activity whilst having insignificant or no noteworthy side-effects at efficacious doses. PCT Application Publication. WO2008095809A1 discloses derivatives of known corticosteroids, containing a NO-releasing moiety which are useful in the treatment of illnesses wherein the known corticosteroid, parent or precursor steroid, is generally applied, with increased benefit in terms of pharmacological profile and fewer or milder side effects than those of the parent corticosteroids.
The NO-releasing derivatives and prodrugs of various therapeutic agents known in the art are in different phases of clinical development and there are reports suggesting that a few of them have been suspended because of some problems (see press reports on naproxcinod and NCX4016 at www.nicox.com). Therefore, there is a clear unmet medical need for new, alternative and better NO-releasing nitrate ester prodrug compounds which can exhibit improved therapeutic properties.
One such class of compounds can be represented by the following generic or Markush structure (IA):

Wherein,
Dx represents a part of a drug or therapeutic agent containing at least one carboxylic acid group which forms a bio-cleavable ester bond with the specified linker and such drug or therapeutic agent is selected from the group consisting of non-steroidal anti-inflammatory, analgesic and antipyretic drugs such as aspirin, diclofenac, naproxen and the like, COX-2 inhibitors, angiotensin-II receptor blockers such as sartans (i.e., losartan, valsatan, candesartan, telmisartan, eprosartan and olmesartan), ACE inhibitors such as captopril, enalapril and the like, beta (β)-blockers such as timolol, atenolol and the like, HMG-CoA reductase inhibitors (cholesterol-reducing agents) such as statins (i.e., fluvastatin, pravastatin, cerivastatin, atorvastatin and rosuvastatin), antiulcerative agents such as misoprostol acid and so on among others;
Xz independently represents at each occurrence a linear or branched alkylene C1-C20 preferably alkylene C1-C10, yet preferably alkylene C1-C6, yet preferably alkylene C2-C10, substituted alkylene C1-C20, substituted alkylene C1-C10, cycloalkylene C3-C7, cycloalkylene C5-C7, optionally substituted cycloalkylene C3-C7 or C5-C7, or [C(Ra)(Rb)]m,
Wherein,
m=1-20, preferably 1-10, yet preferably 1-6 or 2-10 or 2-5;
Ra and Rb at each occurrence are independently a hydrogen, substituted or unsubstituted straight or branched alkyl C1-C20, preferably alkyl C1-C10, or yet preferably alkyl C1-C6 or
Ra and Rb taken together with the carbon atom to which they are attached form a cycloalkyl group, and so on among others;
The above Markush formula (IA) is deduced from the following 20 relevant patent applications.    1. WO2007054451 (Nicox S. A., Fr.).    2. CN101053662 (Jiangsu Wuzhong Suyao Drugs Development Co., Ltd., Peop. Rep. China).    3. WO2005070868 (Merck Frosst Canada & Co., Can.).    4. WO2005030224 (Nicox S. A., Fr.).    5. WO2005011646; Family: AU2004260830 (Nicox S. A., Fr.).    6. WO2004035042, Family: AU2003269774 (Astrazeneca UK Limited, UK).    7. WO2004004648, Family: CA2491127 (Nitromed, Inc., USA).    8. WO2003094923, Family: AU2003236636 (Scaramuzzino, Giovanni), WO2003084550, Family: AU2003224002 (Nicox S. A., Fr.).    9. EP1219306, Family: AU2002219225 (Nicox S. A., Fr.).    10. WO9821193, Family: CA2272063 (Nicox S. A., Fr.; Del Soldato, Piero).    11. WO9809948, Family: EP931065 (Nicox S. A., Fr.).    12. WO9716405, Family: EP871606 (Nicox S. A., Fr.).    13. WO9530641, Family: EP759899 (Nicox Ltd., Ire.).    14. WO2007088123, Family: AU2007211508 (Nicox S. A., Fr.).    15. CN1966484 (Beijing Meibeita Pharmaceutical Research Co., Ltd., Peop. Rep. China).    16. WO2004020384, Family: EP1532098 (Nicox S. A., Fr.).    17. WO2001010814, Family: EP1200386 (Nicox S. A., Fr.).    18. WO2009000592, Family: EP2164484 (Nicox S. A., Fr.).    19. WO2004105754, Family: US7166638 (Nicox S. A., Fr.).    20. WO9858910, Family: EP989972 (Nicox S. A., Fr.).
We now report a small set of compounds of formula (I) which possesses surprising and unexpected properties when compared with compounds of formula (IA).

Wherein,
Dx is a part of a drug/therapeutic agent containing at least one carboxylic acid group [i.e., DxCO2H] which is covalently bonded to the specified linker “C(H)(Ry)” via a bio-cleavable ester linkage;
Ry is an alkyl C1-C6 or cycloalkyl C3-C7; preferably alkyl C1-C4; yet preferably alkyl C1-C2; yet most preferably Ry is methyl (i.e., CH3);
ONO2 (a nitrooxy) group is covalently bonded to the other side of the linker;
and all its geometrical and stereoisomeric forms and pharmaceutically acceptable salts thereof.
The compounds of the present invention represented by formula (I) are generically covered within the scope of some of the patents or patent applications listed above.
Obviously, the Markush formula (IA), with so many variables, would encompass several thousand or even millions of possible compounds including the compounds of formula (I) of this invention. However, none of the above mentioned prior art documents specifically disclosed or claimed any of the possible compounds of this invention that are represented specifically by the formula (I).
A characteristic and unique structural feature of the specific set of compounds of formula (I) of the invention (i.e., representing a species) when compared to those of the compounds of formula IA (i.e., representing a genus) is the presence of a unique “acyl-acetal” type linkage represented by “—C(═O)—O—C(H)(Ry)-O—” group, which is a “hybrid” form of an ester and an acetal group. This characteristic and unique structural feature possibly imparts hitherto undisclosed properties to the compounds containing this “acyl-acetal” type linkage which are essentially the compounds of this invention specifically represented by the formula (I).
Some of the characteristic properties exhibited by these unique set of compounds include:    1. The compounds of formula (I) are the only kind of nitric oxide releasing ester prodrugs of carboxyl-containing drugs that encompass the unique “acyl-acetal” type structural feature.    2. Upon incubation in simulated gastric and/or intestinal fluid/s, the compounds of formula (I) readily released significant amounts of parent drugs (including aspirin!). It is well known to the people skilled in the art that it has been a very difficult task to design a true ester prodrug of aspirin due to the presence of a very labile acetyl group which undergoes preferential hydrolysis by plasma esterases.
Consequently, a vast majority of ester prodrugs of aspirin turn out be prodrugs of salicylic acid. However, in case of aspirin, the promising NO-aspirin prodrug I-D1-R1 (i.e. Dx=D1=aspirin; Ry=R1=CH3) of the present invention was seen to act as a true prodrug of aspirin, when tested for its capability to release aspirin in Simulated Gastric Fluid (SGF) (FIG. 8) and Simulated Intestinal Fluid (SIF) (FIG. 9). The prodrug I-D1-R1 was evaluated at a concentration of either 100 μM or 1 mM in SGF (aspirin was co-evaluated as a positive control under the same experimental conditions, at equimolar doses) and has shown dose dependent decrease/increase in the amount of aspirin released. In SIF also, the prodrug I-D1-R1 released significant amount of aspirin at 1 mM concentration. However, although the aspirin release increased in a dose-dependent manner, it was significantly less than that of aspirin standard at equimolar doses. In SIF, with its pH in the range of ˜6-7, a certain percentage of the prodrug preferentially underwent de-acetylation to give salicylic acid intermediate which further degraded to salicylic acid.
Interestingly, the behaviour of NO-aspirin (i.e. Dx=D1=aspirin) prodrugs of formula (I) was seen to be significantly different from the analogous compounds of formula (IA) [(i.e., with the same molecular formula and molecular weight but with different structural features; i.e., structures I-D1-R1 and II-D1-X2, respectively). When these two compounds were incubated simultaneously in SGF, it was observed that only the compound of formula (I), i.e., I-D1-R1, of the present invention, released quantitative amounts of the parent drug aspirin whereas the compound of formula (IA) i.e., II-D1-X2, quickly decomposed into an unknown metabolite, without releasing even traces of aspirin. Additionally, another analogous NO-aspirin compound NCX-4016 of formula (IA) that had reached phase II clinical trials (structure shown below) remained intact (no release of parent drug, aspirin) when incubated in SGF under identical conditions (Table 1).

TABLE 1Stability study of NO-aspirin prodrugs in SGFI-D1-R1II-D1-X2II-D1-X1 [NCX-4016](Ry = R1 = CH3)[Xz = X2 = CH2CH2][Xz = X1 = m-C6H4CH2]Time% of Prodrug% of Aspirin% of Prodrug% of Aspirin% of Prodrug% of AspirinPointremainingReleasedremainingReleasedremainingReleased(min)(μM)(μM)(μM)(μM)(μM)(μM)0910The prodrugAspirin100.00Aspirin58218underwentrelease was100.00release was105644quicknot observed100.00not observed30397decomposition100.00600100into an100.001200100unknown100.00metabolitet1/2<15 min~1.5 min—
In case of naproxen series also, the behaviour of NO-naproxen (i.e. Dx=D2=naproxen) prodrugs of formula (I) was seen to be significantly different from the analogous compounds of formula (IA) (i.e., with the same molecular formula and molecular weight but with different structural features; See structures I-D2-R1 and II-D2-X2, shown below); when incubated simultaneously in SGF, it was observed that only the compound of formula (I) of the present invention i.e., I-D2-R1 released quantitative amounts of the parent drug, naproxen whereas the compound of formula (IA) i.e., II-D2-X2 remained intact (no release of parent drug naproxen) under identical conditions (Table 2).

TABLE 2Stability of prodrugs I-D2-R1 and II-D2-X2 in SGFI-D2-R1 (Ry = R1 = CH3)II-D2-X2 [Xz = X2 = (CH2)2]TimeI-D2-R1NaproxenII-D2-X2Naproxen(mins)Remaining (%) Released (%) Remaining (%)Released (%)0100.000.00100.000.00584.6815.3299.680.321074.5825.4299.790.211564.5835.4299.680.323032.7967.2199.750.25600.00100.0099.670.331200.00100.0099.390.611800.00100.0099.190.81t1/220-25 minNA
Even the higher homologue pairs of naproxen prodrugs of formula (I) and formula (IA) i.e., I-D2-R2 vs II-D2-X3 and I-D2-R3 vs II-D2-X4 behaved in a similar fashion, when incubated simultaneously in SGF. Thus only compounds of formula (I) i.e., I-D2-R2 and I-D2-R3 released naproxen (Tables 3 and 4, respectively).

TABLE 3Stability of prodrugs I-D2-R2 and II-D2-X3 in SGFI-D2-R1 (Ry = R2 = CH3CH2)II-D2-X3 [Xz = X3 = (CH2)3]TimeI-D2-R2NaproxenII-D2-X3Naproxen(mins)Remaining (%) Released (%) Remaining (%)Released (%)01000.00100.000.00594.335.67100.000.001090.439.57100.000.001574.2725.73100.000.003062.1537.85100.000.006051.0148.99100.000.0012039.0860.92100.000.001806.7793.23100.000.00t1/2~1 hNA

TABLE 4Stability of prodrugs I-D2-R3 and II-D2-X4 in SGFI-D2-R3 II-D2-X4 (Ry = R3 = CH3CH2CH2)[Xz = X4 = (CH2)4]I-D2-R3NaproxenII-D2-X4NaproxenTimeRemaining Remaining Remaining Remaining (mins)(%)(%)(%)(%)01000.00100.000.00596.353.65100.000.001093.616.39100.000.001583.2116.79100.000.003082.7317.27100.000.006075.4824.52100.000.0012057.5542.45100.000.0018039.0061.00100.000.00t1/2~2.5 hNA
A still higher homologue of naproxen prodrug of formula (I), i.e., I-D2-R4, also released appreciable amounts of the parent drug naproxen when incubated in SGF, as shown below (Table 5).

TABLE 5Stability of prodrug I-D2-R4 in SGFI-D2-R4 (Ry = R4 = CH3CH2CH2CH2)TimeI-D2-R4Naproxen(mins)Remaining (%)Released (%)01000.00597.742.261096.183.821587.4112.593090.539.476088.8711.1312079.9720.0318071.0428.96t1/2>3 h
From the above data, it is obvious that the compounds (prodrugs of naproxen) of formula (I) release decreasing amounts of parent drug naproxen with increasing alkyl chain length of Ry (probably due to solubility issues associated with increased hydrophobicity of longer alkyl chains). Thus, the half-lives (t1/2) of prodrugs of naproxen of formula (I) follow the pattern: I-D2-R1 (t1/2=˜20-25 min)<I-D2-R2 (t1/2=˜1 h)<I-D2-R3 (t1/2=˜2.5 h)<I-D2-R4 (t1/2=>3 h).
Similarly, I-D3-R1, which is the nitric oxide releasing prodrug of chlorambucil of formula (I) (i.e., Dx=D3=chlorambucil, Ry=R1=CH3) also released its parent drug chlorambucil quantitatively when incubated in SGF as shown below (See Table 6 and FIG. 12).

TABLE 6Stability of prodrug I-D3-R1 in SGFI-D3-R1 (Ry = R1 = CH3)TimeI-D3-R1Chlorambucil(mins)Remaining (%)Released (%)092.127.88520.4979.511013.8486.16158.5991.41300.00100.00600.00100.001200.00100.001800.00100.00t1/2<5 min
Interestingly, the chlorambucil prodrug I-D3-R1, which is the lowest carbon homologue among the chlorambucil prodrugs of formula (I), decomposed in SGF to give 100% of the parent drug chlorambucil, with a half-life of less than 5 minutes (Table 6).    3. The compounds of formula (I) exhibited nearly similar or superior oral bioavailability and efficacy as compared to those of respective parent drugs in rats (See FIGS. 1, 2, 3 and Table 7).    4. Although the compounds of formula (I) at equimolar doses exhibited nearly similar or superior oral bioavailability and efficacy as compared to those of their respective parent drugs, they did not cause any significant drug-induced gastric lesions and/or bleeding. However, their respective parent drugs at equimolar doses caused significant drug-induced gastric lesions and/or bleeding (See FIGS. 5 and 6).    5. The process for making the compounds of formula (I) differs significantly when compared to the reported processes for making the compounds of formula (IA).
For example, the most frequently used process for making the compounds of formula (IA) involves the steps, as shown in the Chart 1A:

Step 1: Conversion of the drug or therapeutic agent containing carboxylic acid group (Dx-CO2H) to its active acid chloride Dx-C(═O)Cl by reacting with thionyl chloride or oxalyl chloride in presence of catalytic amount of DMF;
Step 2: Conversion of diol HO-Xz-OH for example 1,2-Ethanediol or 1,3-Propanediol or 1,4-Butanediol (wherein Xz is as defined above), into its mono bromide derivative HO-Xz-Br, by known methods for example by treating with carbon tetrabromide and triphenylphosphine in a solvent such as DCM;
Step 3: Conversion of monobromide HO-Xz-Br from Step 3 into the corresponding mononitrate HO-Xz-ONO2 by treating with silver nitrate in acetonitrile;
Step 4: Reaction of acid chloride from Step 1 with the mononitrate from Step 3 in the presence of a suitable base such as triethylamine in a suitable solvent such as DCM to yield the compound of formula (IA).
In contrary, the process for making the compounds of formula (I) involves significantly different steps as shown in Schemes 1 and 2.
For clarity, a plausible mechanism for the formation of the compounds of formula (I) is shown below:

It would be understood by a person skilled in the art that in the compounds of formula (I), the “CO” group adjacent to Dx is derived from the carboxyl group of the drug (i.e., Dx-CO2H) as shown in chart 1B.
The above mentioned characteristic properties of the unique compounds of the present invention represented specifically by the formula (I) [i.e., representing a specific species comprising the compounds of formula (I)] are neither disclosed specifically in the prior art [i.e., representing the whole genus comprising the compounds of formula (IA)] nor obvious to those skilled in the art to which this invention relates. The unique structural features and characteristic properties of the compounds of formula (I) therefore constitute or impart both “novelty and inventive features” to these potentially useful compounds.